This invention relates to a certain class of compounds, and the pharmaceutically acceptable salts, solvates and prodrugs thereof, which inhibit Procollagen C-proteinase (xe2x80x9cPCPxe2x80x9d). These compounds are useful in the treatment of mammals having conditions alleviable by inhibition of PCP. Especially of interest is an antiscarring treatment for wounds.
Fibrotic tissues, including dermal scars, are characterised by excessive accumulation of extracellular matrix, mainly collagen type I. It is thought that inhibition of collagen deposition will reduce formation of scar tissue. Collagen is secreted as the precursor, procollagen, which is transformed into the insoluble collagen by cleavage of the C-terminal propeptide by PCP. PCP is a zinc-dependent metalloprotease which is secreted from TGF-xcex2-activated fibroblasts belonging to the subfamily of astacin-like proteases and able to cleave the C-terminal peptide of types I, II and III procollagens. Furthermore, data suggest that PCP activates lysyl oxidase, an enzyme essential for the formation of covalent cross-links which stabilise the fibrous form of collagen. Therefore, inhibition of PCP may not only reduce collagen deposition but may also make collagen more accessible for degradation.
Collagen is integral to, among other things, the proper formation of connective tissue. Thus, the over- or under-production of collagen or the production of abnormal collagen (including incorrectly processed collagen) has been linked with numerous connective tissue diseases and disorders. Mounting evidence suggests that PCP is an essential key enzyme for the proper maturation of collagen (see for example International Patent Application publication number WO 97/05865).
At present more than nineteen types of collagens have been identified. These collagens, including fibrillar collagen types I, II, III are synthesized as procollagen precursor molecules which contain amino- and carboxy-terminal peptide extensions. These peptide extensions, referred to as xe2x80x9cpro-regions,xe2x80x9d are designated as N- and C-propeptides, respectively.
The pro-regions are typically cleaved upon secretion of the procollagen triple helical precursor molecule from the cell to yield a mature triple helical collagen molecule. Upon cleavage, the xe2x80x9cmaturexe2x80x9d collagen molecule is capable of association, for example, into highly structured collagen fibers. See e.g., Fessler and Fessler, 1978, Annu. Rev. Biochem. 47:129-162; Bornstein and Traub, 1979, in: The Proteins (eds. Neurath, H. and Hill, R. H.), Academic Press, New York, pp. 412-632; Kivirikko et al., 1984, in: Extracellur Matrix Biochemistry (eds. Piez, K. A. and Reddi. A. H.), Elsevier Science Publishing Co., Inc., New York, pp. 83-118; Prockop and Kivirikko, 1984, N. Engl. J. Med. 311:376-383; Kuhn, 1987, in: Structure and Function of Collagen Types (eds. Mayne, R. and Burgeson, R. E.), Academic Press, Inc., Orlando, Fla., pp. 1-42.
An array of conditions has been associated with the inappropriate or unregulated production of collagen, including pathological fibrosis or scarring, including endocardial sclerosis, idiopathic interstitial fibrosis, interstitial pulmonary fibrosis, perimuscular fibrosis, Symmers"" fibrosis, pericentral fibrosis, hepatitis, dermatofibroma, cirrhosis such as binary cirrhosis and alcoholic cirrhosis, acute pulmonary fibrosis, idiopathic pulmonary fibrosis, acute respiratory distress syndrome, kidney fibrosis/glomerulonephritis, kidney fibrosis/diabetic nephropathy, scleroderma/systemic, scleroderma/local, keloids, hypertrophic scars, severe joint adhesions/arthritis, myelofibrosis, corneal scarring, cystic fibrosis, muscular dystrophy (duchenne""s), cardiac fibrosis, muscular fibrosis/retinal separation, esophageal stricture and Pyronie""s disease. Further fibrotic disorders may be induced or initiated by surgery, including scar revision/plastic surgeries, glaucoma, cataract fibrosis, corneal scarring, joint adhesions, graft vs. host disease, tendon surgery, nerve entrapment, dupuytren""s contracture, OB/GYN adhesions/fibrosis, pelvic adhesions, peridural fibrosis and restenosis. Other conditions where collagen plays a key role include burns. Fibrosis of lung tissue is also observed in patients suffering from chronic obstructive airways disease (COAD) and asthma. One strategy for the treatment of these diseases and conditions is to inhibit the overproduction and/or deposition and/or unregulation of collagen. Thus, identification and isolation of molecules which control, inhibit and/or modulate the production and deposition of collagen are of major medical interest.
Recent evidence suggests that PCP is the essential key enzyme that catalyzes the cleavage of the Procollagen C-propeptide. This has been demonstrated in fibrillar collagens, including type I, type II, and type III collagen.
PCP was first observed in the culture media of human and mouse fibroblasts (Goldberg et al., 1975, Cell 4:45-50; Kessler and Goldberg, 1978, Anal. Biochem. 86:463-469), and chick tendon fibroblasts (Duskin et al., 1978, Arch. Biochem. Biophys. 185:326-332; Leung et al., 1979, J. Biol. Chem. 254:224-232). An acidic proteinase which removes the C-terminal propeptides from type I procollagen has also been identified (Davidson et al., 1979, Eur. J. Biochem. 100:551).
A partially purified protein having PCP activity was obtained from chick calvaria in 1982. Njieha et al., 1982, Biochemistry 23:757-764. In 1985, chicken PCP was isolated, purified and characterized from conditioned media of chick embryo tendons. Hojima et al., 1985, J. Biol. Chem. 260:15996-16003. Murine PCP has been subsequently purified from media of cultured mouse fibroblasts. Kessler et al., 1986, Collagen Relat. Res. 6:249-266; Kessler and Adar, 1989, Eur. J. Biochem. 186:115-121. Finally, the cDNA encoding human PCP has been identified, as set forth in the above-referenced articles and references disclosed therein.
Experiments conducted with these purified forms of chick and mouse PCP have indicated that the enzyme is instrumental in the formation of functional collagen fibers. Fertala et al., 1994, J. Biol. Chem. 269:11584.
As a consequence of the enzyme""s apparent importance to collagen production, scientists have identified a number of PCP inhibitors. See, e.g., Hojima et al., supra. For example, several metal chelators have demonstrated activity as PCP inhibitors. Likewise, chymostatin and pepstatin A were found to be relatively strong inhibitors of PCP. Additionally, xcex12-Macroglobulin, ovostatin, and fetal bovine serum appear to at least partially inhibit PCP activity.
Dithiothreitol, SDS, concanavalin A, Zn2+, Cu2+, and Cd2+ are similarly reported to be inhibitory at low concentrations. Likewise, some reducing agents, several amino acids, phosphate, and ammonium sulfate were inhibitory at concentrations of 1-10 mM. Further, the enzyme was shown to be inhibited by the basic amino acids lysine and arginine (Leung et al., supra; Ryhxc3xa4nen et al., 1982, Arch. Biochem. Biophys. 215:230-235). Finally, high concentrations of NaCl or Tris-HCl buffer were found to inhibit PCP""s activity. For example, it is reported that, with 0.2, 0.3, and 0.5M NaCl, the activity of PCP was reduced 66, 38, and 25%, respectively, of that observed with the standard assay concentration of 0.15M. Tris-HCl buffer in a concentration of 0.2-0.5M markedly inhibited activity (Hojima et al., supra). PCP activity and its inhibition have been determined using a wide array of assays. See e.g., Kessler and Goldberg, 1978, Anal. Biochem. 86:463; Njieha et al., 1982, Biochemistry 21:757-764. As articulated in numerous publications, the enzyme is difficult to isolate by conventional biochemical means and the identity of the cDNA sequence encoding such enzyme was not known until reported in the above referenced and related patent applications.
In view of its essential role in the formation and maturation of collagen, PCP appears to be an ideal target for the treatment of disorders associated with the inappropriate or unregulated production and maturation of collagen.
Matrix metalloproteases (MMPs) constitute a family of structurally similar zinc-containing metalloproteases, which are involved in the remodelling, repair and degradation of extracellular matrix proteins, both as part of normal physiological processes and in pathological conditions.
Another important function of certain MMPs is to activate other enzymes, including other MMPs, by cleaving the pro-domain from their protease domain. Thus, certain MMPs act to regulate the activities of other MMPs, so that over-production in one MMP may lead to excessive proteolysis of extracellular matrix by another, e.g. MMP-14 activates pro-MMP-2.
During the healing of normal and chronic wounds, MMP-1 is expressed by migrating keratinocytes at the wound edges (U. K. Saarialho-Kere, S. O. Kovacs, A. P. Pentland, J. Clin. Invest. 1993, 92, 2858-66). There is evidence which suggests MMP-1 is required for keratinocyte migration on a collagen type I matrix in vitro, and is completely inhibited by the presence of the non-selective MMP inhibitor SC44463 ((N-4-hydroxy)-N1-[(1S)-2-(4-methoxyphenyl)methyl-1-((1R)-methylamino)carbonyl)]-(2R)-2-(2-methylpropyl)butanediamide) (B. K. Pilcher, J. A. Dumin, B. D. Sudbeck, S. M. Krane, H. G. Welgus, W. C. Parks, J. Cell Biol., 1997, 137, 1-13). Keratinocyte migration in vivo is essential for effective wound healing to occur.
MMP-2 and MMP-9 appear to play important roles in wound healing during the extended remodelling phase and the onset of re-epithelialisation, respectively (M. S. Agren, Brit. J. Dermatology, 1994, 131, 634-40; T. Salo, M. Mxc3xa4kxc3xa4nen, M. Kylmxc3xa4niemi, Lab. Invest., 1994, 70, 176-82). The potent, non-selective MMP inhibitor BB94 ((2S,3R)-5-methyl-3-{[(1S)-1-(methylcarbamoyl)-2-phenylethyl]carbamoyl}-2-[(2-thienylthio)methyl]hexanohydroxamic acid, batimastat) inhibits endothelial cell invasion of basement membrane, thereby inhibiting angiogenesis (G. Tarboletti, A. Garofalo, D. Belotti, T. Drudis, P. Borsotti, E. Scanziani, P. D. Brown, R. Giavazzi, J. Natl. Cancer Inst., 1995, 87, 293-8). There is evidence that this process requires active MMP-2 and/or 9.
Thus PCP inhibitors which significantly inhibit MMPs 1 and/or 2 and/or 9 would be expected to impair wound healing. MMP-14 is responsible for the activation of MMP-2, and thus inhibition of MMP-14 might also result in impaired wound healing.
For recent reviews of MMPs, see Zask et al., Current Pharmaceutical Design, 1996, 2, 624-661; Beckett, Exp. Opin. Ther. Patents, 1996, 6, 1305-1315; and Beckett et al., Drug Discovery Today, vol. 1 (no.1), 1996, 16-26.
Alternative names for various MMPs and substrates acted on by these are shown in the table below (Zask et al., supra).
International Patent Applications PCT/IB00/01855 (published as WO 01/47901) and PCT/IB01/02360 (filed on 7th Dec. 2001), and foreign equivalents thereof, describe various 3-heterocyclylpropanohydroxamic acid PCP inhibitors. The teachings of these documents are herein incorporated by reference in their entirety.
The identification of effective compounds which specifically inhibit the activity of PCP to regulate and modulate abnormal or inappropriate collagen production is therefore desirable and the object of this invention.
According to one aspect of the present invention, there are provided compounds of formula (I): 
wherein [HET] is a divalent heterocyclic moiety selected from: 
X is C1-6 alkylene or C2-6 alkenylene, each of which is optionally substituted by one or more fluorine atoms; R is aryl, C5-8 cycloalkenyl or C3-8 cycloalkyl optionally substituted by one or more fluorine atoms;
Z is H or C1-4 alkyl;
Y is a mono- or bicyclic unsaturated ring system containing from 5 to 10 ring atoms, of which up to 4 which are hetero-atoms independently selected from N, O and S, and which ring system is optionally substituted by one or more substituents independently selected from xe2x95x90O, C1-4 alkyl, C1-4 alkoxy, NR1R2, SO2NR1R2, CO2R1, CONR1R2, CH2CO2R1, NR1CO2R2, NR1SO2R2 or het1;
R1 and R2 are each independently selected from H and C1-4 alkyl optionally substituted by C1-4 alkoxy;
het1 is a N-linked 4- to 6-membered mono- or bicyclic heterocycle optionally containing 1 or 2 further hetero ring atoms independently selected from N and O, which heterocycle is optionally substituted by one or more substituents independently selected from xe2x95x90O, C1-4 alkyl, C1-4 alkoxy, NR1R2, SO2NR1R2, CO2R1, CONR1R2, CH2CO2R1, NR1CO2R2, NR1SO2R2 or het2, het2 is a N-linked 4- to 6-membered mono- or bicyclic heterocycle optionally containing 1 or 2 further hetero ring atoms independently selected from N and O;
xe2x80x9carylxe2x80x9d is phenyl optionally substituted by one or more substituents independently selected from R1, OH, SO2(C1-4 alkyl) and/or C(O)p(C1-4 alkyl) groups;
and the pharmaceutically acceptable salts, solvates (including hydrates) and prodrugs thereof.
xe2x80x9cAlkylxe2x80x9d, xe2x80x9calkylenexe2x80x9d, xe2x80x9calkoxyxe2x80x9d, xe2x80x9calkanoylxe2x80x9d, and xe2x80x9calkenylenexe2x80x9d groups, including groups incorporating said moieties, may be straight chain or branched where the number of carbon atoms allows.
Halogen is taken to mean fluorine, chlorine, bromine or iodine.
Pharmaceutically-acceptable salts are well known to those skilled in the art, and for example include those mentioned in the art cited above, and by Berge et al, in J. Pharm. Sci., 66, 1-19 (1977). Suitable acid addition salts are formed from acids which form non-toxic salts and include the hydrochloride, hydrobromide, hydroiodide, nitrate, sulphate, bisulphate, phosphate, hydrogenphosphate, acetate, trifluoroacetate, gluconate, lactate, salicylate, citrate, tartrate, ascorbate, succinate, maleate, fumarate, gluconate, formate, benzoate, methanesulphonate, ethanesulphonate, benzenesulphonate, pamoate, carasylate, and p-toluenesulphonate salts.
Pharmaceutically acceptable base addition salts are well known to those skilled in the art, and for example include those mentioned in the art cited above, and can be formed from bases which form non-toxic salts and include the aluminium, calcium, lithium, magnesium, potassium, sodium and zinc salts, and salts of non-toxic amines such as diethanolamine.
Certain of the compounds of formula (I) may exist in one or more zwitterionic forms. Certain of the compounds of formula (I) may exist in one or more tautomeric forms. Certain of the compounds of formula (I), their salts, solvates, prodrugs, etc. may exist in one or more polymorphic forms. It is to be understood that the compounds of formula (I) include all such zwitterions, tautomers and polymorphs.
The compounds of formula (I), their salts, hydrates, prodrugs etc. can exhibit isotopic variation, e.g. forms with enriched 2H, 13C, 14C, 15N, 18O, etc. may be prepared, for example by suitable variation of the synthetic methods described herein using methods and reagents known in the art or routine modification thereof. All such isotopic variants are included in the scope of the invention.
Prodrug moieties are well-known to those skilled in the art (see for example the article by H Feres, in Drugs of Today, vol 19, no.9 (1983) pp.499-538, especially section Al), and for example include those specifically mentioned in A. A. Sinkula""s article in Annual Reports in Medicinal Chemistry, vol 10, chapter 31, pp.306-326, herein incorporated by reference, and the references therein. Specific prodrug moieties which may be specifically mentioned are aliphatic-aromatic, carbonate, phosphate and carboxylic esters, carbamates, peptides, glycoside, acetals and ketals, tetrahydropyranyl and silyl ethers. Such prodrug moieties can be cleaved in situ, e.g. are hydrolysable in physiological conditions, to give compounds of formula (I).
Certain of the compounds of the formula (I) may exist as geometric isomers. The compounds of the formula (I) may possess one or more asymmetric centers, apart from the specified centers in formula (I), and so exist in two or more stereoisomeric forms. The present invention includes all the individual stereoisomers and geometric isomers of the compounds of formula (I) and mixtures thereof.
Preferably the compounds of formula (I) have the following stereochemistry (IA): 
Preferably X is a linear C2-6 alkylene moiety optionally substituted by one or more fluorine atoms. More preferably X is propylene.
Preferably [HET] is 
Preferably R is C3-8 cycloalkyl optionally substituted by one or more fluorine atoms. More preferably R is cyclobutyl, cyclopentyl or cyclohexyl optionally substituted by one or more fluorine atoms. Most preferably R is cyclohexyl.
Preferably Y is a 5- or 6-membered unsaturated ring containing 0, 1 or 2 ring hetero-atoms independently selected from N and O,
and which ring is optionally substituted by one or more substituents independently selected from xe2x95x90O, C1-4 alkyl, C1-4 alkoxy, NR1R2, SO2NR1R2, CO2R1, CONR1R2, CH2CO2R1, NR1CO2R2, NR1SO2R2 and het1.
More preferably Y is a 5- or 6-membered unsaturated ring containing 0, 1 or 2 ring hetero-atoms independently selected from N and O, and which ring is optionally substituted by one or more substituents independently selected from xe2x95x90O, CONH2, CONHCH3, CON(CH3)2, CH3, NH2, CO2C2H5, CO2H, CH2CO2CH3, NHCH3, NHC2H5, N(CH3)2, morpholino, imidazol-1-yl, methoxy, ethoxy and NHSO2CH3.
Yet more preferably Y is 5-carbamoylpyridin-3-yl, uracil-5-yl, pyridin-4-yl, 6-methylpyridin-3-yl, 5-carboxypyridin-3-yl, 6-methoxycarbonylmethylpyridin-3-yl, 6-(4-methylpiperazino)pyridin-3-yl, 6-(3-dimethylaminoazetidino)pyridin-3-yl, 6-(3-morpholinoazetidino)pyridin-3-yl, 4-carboxypyridin-2-yl, 6-dimethylaminopyridin-3-yl, 6-(imidazol-1-yl)pyrazin-2-yl, 3-morpholinopyrazin-2-yl, 3-pyrrolidinopyrazin-2-yl, 3-dimethylaminopyrazin-2-yl, 3-methylaminopyrazin-2-yl, 3-methylsulphonamidophenyl, 3-carboxyphenyl or 6-ethoxypyrazin-2-yl. Most preferably Y is uracil-5-yl or 4-carboxypyridin-2-yl.
A preferred group of compound of formula (I) are those wherein each substituent is as specified in the Examples below.
Another preferred group are the compounds of the Examples below and the salts, solvates and prodrugs thereof. Especially preferred are the compounds of Examples 1 and 25 and the salts, solvates and prodrugs thereof.
A further aspect of the invention is a PCP inhibitor of formula (I) which is selective against MMP-1 and/or MMP-2 and/or MMP-9 and/or MMP-14.
A further aspect of the invention is the use of a PCP inhibitor of formula (I) which is selective against MMP-1 and/or MMP-2 and/or MMP-9 and/or MMP-14 in medicine.
Further related to this aspect of the invention is the use of a PCP inhibitor of formula (I) which is selective against MMP-1 and/or MMP-2 and/or MMP-9 and/or MMP-14 in the manufacture of an antiscarring medicament.
Further related to this aspect of the invention is a method of treating a condition mediated by PCP and in which MMP-1 and/or MMP-2 and/or MMP-9 and/or MMP-14 have a beneficial effect, with an effective amount of PCP inhibitor of formula (I) which is selective against MMP-1 and/or MMP-2 and/or MMP-9 and/or MMP-14, an example of such a condition being a wound.
Preferably the PCP inhibitor of formula (I) mentioned in this aspect of the invention is selective against at least MMP-1, MMP-2 and MMP-9. Most preferably the said PCP inhibitor of formula (I) is selective against MMP-1, MMP-2, MMP-9, and MMP-14.
Preferably the selective PCP inhibitor of formula (I) has an IC50 vs. PCP of 0.5 xcexcM or lower, and selectivities vs. MMP-2 and MMP-9 of at least 30-fold, in the tests described herein.
Preferably the selective PCP inhibitor of formula (I) has an IC50 vs. PCP of 0.1M or lower, and selectivities vs. MMP-1, MMP-2, MMP-9 and MMP-14 of at least 300-fold, in the tests described herein.
Another aspect of the invention is a substance of formula (I) described herein, including the salts, solvates and prodrugs thereof, for use in medicine.
Another aspect of the invention is a substance of formula (I) described herein, including the salts, solvates and prodrugs thereof, for use in treating a PCP-mediated condition, such as in an antiscarring medicament.
Another aspect of the invention is the use of the substances of formula (I) described herein, including the salts, solvates and prodrugs thereof, in the manufacture of a medicament for treatment of a PCP-mediated condition (e.g. an antiscarring medicament).
Another aspect of the invention is a pharmaceutical composition comprising a compound of formula (I), salts thereof, solvates thereof and/or prodrugs thereof, and a pharmaceutically acceptable diluent, carrier or adjuvant.
Another aspect of the invention is the combination of a compound of formula (I), or a salt, solvate or prodrug thereof, with another material useful in treating wounds, such as:
(i) a growth factor such as TGF-xcex2-3 (Renovo), IGF-1 (Genentech), IGF-1 complex (Celtrix), KGF-2 or FGF-10 (Sumitomo), DWP-401/EGF (Daewoong) or SNK-863 (Sanwa Kagaku Kenkyusho);
(ii) a growth factor agonist such as Noggin (Regeneron);
(iii) a growth factor antibody/antisense material, such as those to: TGF-xcex2-1 or 2 (Renovo, CaT), PDGF (11 Yang) or CTGF (Fibrogen);
(iv) a hormone such as DHEAS (Pharmadigm), ConXn/Relaxin (Connetics);
(v) an antibody to adhesion compounds such as ICAM-1 (Boehringer);
(vi) a MMP beneficial to healing of wounds, such as Collagenase ABC (BioSpecifics);
(vii) a barrier such as ADCON (Gliatech);
(viii) skin products such as artificial skin systems such as those based on DermaGraft (Advanced Tissue Sciences Inc.), INTEGRA Artificial Skin (Integra Life Sciences Holding Corp.), cell cultures such as Apligraf/Graftskin (Novartis), those developed by Cell Genesys Inc., AlloDerm (LifeCell) or matrix formulation products such as Argidene gel (Telios Pharmaceuticals Inc.);
(ix) a uPA inhibitor such as those disclosed in WO 99/01451; and
(x) a MMP-3 inhibitor such as those disclosed in WO99/35124, WO 99/29667 and WO 00/74681.
Yet another aspect of the invention is a method of treatment of a condition mediated by PCP comprising administration of a therapeutically-effective amount of a substance according to the above definitions.
It is to be appreciated that reference to treatment includes prophylaxis as well as the alleviation of established symptoms of PCP-mediated conditions and diseases.
The invention further provides Methods for the production of compounds of the invention, which are described below and in the Examples and Preparations. The skilled man will appreciate that the compounds of the invention could be made by methods other than those specifically described herein, by adaptation of the methods herein described in the sections below and/or adaptation thereof, for example by methods known in the art. Suitable guides to synthesis, functional group transformations, use of protecting groups, etc. are, for example, xe2x80x9cComprehensive Organic Transformationsxe2x80x9d by R C Larock, VCH Publishers Inc. (1989), xe2x80x9cAdvanced Organic Chemistryxe2x80x9d by J March, Wiley Interscience (1985), xe2x80x9cDesigning Organic Synthesisxe2x80x9d by S Warren, Wiley Interscience (1978), xe2x80x9cOrganic Synthesisxe2x80x94The Disconnection Approachxe2x80x9d by S Warren, Wiley Interscience (1982), xe2x80x9cGuidebook to Organic Synthesisxe2x80x9d by R K Mackie and D M Smith, Longman (1982), xe2x80x9cProtective Groups in Organic Synthesisxe2x80x9d by T W Greene and P G M Wuts, John Wiley and Sons Inc. (1999), and P J Kocienski, in xe2x80x9cProtecting Groupsxe2x80x9d, Georg Thieme Verlag (1994), and any updated versions of said standard works.
In the Methods below, unless otherwise specified, the substituents are as defined above with reference to the compounds of formula (I) above.